Inhibition of noninactivating Na channels of mammalian optic nerve as a means of preventing optic nerve degeneration associated with glaucoma

ABSTRACT

A method and composition for altering a plausible sequence of pathological events in retinal ganglion cells associated with glaucoma, the sequence including membrane depolarization, influx of millimolar amounts of Na +  via non-inactivating Na +  channels, and the lethal elevation of cell Ca 2+  due to reversal of the Na + /Ca 2+  exchanger. The method includes blocking, by administration of a selected composition, of associated, non-inactivating Na +  channels in retinal ganglion cells in order to limit Na + /Ca +  exchange in the retinal ganglion cells and prevent buildup of the Ca 2+  level in the retinal ganglion cells to a lethal level. The results in a method of preventing retinal ganglion cell death, associated with glaucoma, by administering to the optic nerve of a mammal, a compound which blocks the non-inactivating sodium ion channels of the optic nerve. Alternately, said invention relates to a method of preventing optic retinal ganglion cell death in a human by administering to the retinal ganglion cells of said human a compound which blocks the non-inactivating sodium ion channel of the retinal ganglion cells.

BACKGROUND OF INVENTION

[0001] 1. Field of the Invention

[0002] The present invention relates to a method of preventing retinalganglion cell death, associated with glaucoma, by administering toretinal ganglion cells of a mammal, a compound which blocks the putativenon-inactivating sodium ion channels of the above cell type.

[0003] 2. Brief Description of the Art

[0004] Glaucoma is an optic neuropathy associated with elevatedintraocular pressures which are too high for normal function of the eye,and results in irreversible loss of visual function. (See for example,Dreyer et al “Elevated glutamate levels in the vitreous body of humanand monkeys with glaucoma”, Arch. Ophthalmology 114:299-305, 1996). Itis estimated in medical science that glaucoma afflicts approximately 2per cent of the population over the age of forty years, and is thereforea serious health problem. Ocular hypertension, i.e. the condition ofelevated intraocular pressure, which has not yet caused irreversibledamage, is believed to represent the earliest phase of glaucoma. Manytherapeutic agents have been devised and discovered in the prior art forthe treatment or amelioration of glaucoma and of the condition ofincreased intraocular pressure which precedes glaucoma.

[0005] Primary open angle glaucoma (POAG) is associated with a rise inintraocular pressure (IOP). This increase in IOP is believed tocontribute to the loss of optic nerve function which ultimately leads toblindness. Reduction of IOP is therefore a crucial component in themanagement of POAG. However, in many individuals lowering of IOP is notsufficient or ineffective in preventing vision loss associated withPOAG.

[0006] It is thought that a novel class of sodium channels residingwithin the optic nerve of the rat are responsible for damage to the ratoptic nerve following anoxia or hypoxia. However, in glaucoma thesequence of pathological events leading to the loss of optic nervefunction, is not known.

[0007] The drugs currently utilized in the treatment of glaucoma includemiotics (e.g., pilocarpine, carbachol, and acetylcholinesteraseinhibitors), sympathomimetrics (e.g., epinephrine anddipivalylepinephrine), beta-blockers (e.g., betaxolol, levobunolol andtimolol), alpha-2 agonists (e.g., para-amino clonidine) and carbonicanhydrase inhibitors (e.g., acetazolamide, methazolamide andethoxzolamide). Miotics and sympathomimetics are believed to lowerintraocular pressure by increasing the outflow of aqueous humor, whilebeta-blockers, alpha-2 agonists and carbonic anhydrase inhibitors arebelieved to lower intraocular pressure by decreasing the formation ofaqueous humor. All five types of drugs have potential side effects.Miotics, such as pilocarpine, can cause blurring of vision and othervisual side effects which may either decrease patient compliance orrequire termination of miotic drug therapy. Carbonic anhydraseinhibitors can also cause serious side effects which affect patientcompliance and/or necessitate withdrawal of the drug therapy. At leastone beta-blocker, timolol, has increasingly become associated withserious pulmonary side effects attributable to its effect on beta-2receptors in pulmonary tissue.

[0008] As a result additional antiglaucoma drugs are being developed,e.g., prostaglandin derivatives, muscarinic antagonists, etc. However,none of the above drugs are designed to directly interact with theretinal ganglion cell and its associated axon.

[0009] Thus, it would be desirable to prevent the loss of ganglion cellbody and axon function, which may be associated with glaucoma by abiological mechanism which does not modulate aqueous humor dynamics andtherefore intraocular pressure. Moreover, it would be desirable to treatthe retinal ganglion cell body and axon of a mammal directly to preventthe destruction thereof by the glaucomatous condition.

SUMMARY OF THE INVENTION

[0010] Surprisingly, it has been discovered in accordance with thepresent invention, that sodium channel blockers which block thenon-inactivating sodium ion channel of the optic nerve of a mammal maybe effective for preventing the loss of retinal ganglion cells when suchsodium channel blockers are administered and applied in a pharmaceuticalcomposition. Accordingly, the present invention relates to a method ofpreventing loss of retinal ganglion cells and their associated axons(optic nerve) function, associated with glaucoma, by systemically ordirectly administering to the eye of a mammal an ophthalmic compositionwhich includes an amount of a sodium channel blocker which is effectiveto block the non-inactivating sodium ion channel of the ganglion cellsof said mammal.

[0011] More specifically, the present invention is directed to a methodfor altering a possible sequence of pathological events in retinalganglion cells that may be associated with glaucomatous opticneuropathy. The sequence includes the pathological depolarization ofretinal ganglion cells, an influx of millimolar amounts of sodium vianon-inactivating sodium channels and a subsequent reversal of thesodium/calcium exchanger. Reversal of the sodium/calcium exchangermediated by both membrane depolarization and increased intracellularsodium causes a toxic buildup of intracellular calcium. The method foraltering this sequence includes a step of blocking associatednon-inactivating sodium channels in retinal ganglion cells in order toprevent reversal of sodium/calcium ion exchange and subsequent buildupof the calcium ion concentration in the retinal ganglion cells to alethal level.

[0012] Specifically, this blocking is achieved by administering to theretinal ganglion cells a pharmaceutical composition having an activeingredient with non-inactivating sodium channel blocking activity.

[0013] Specific examples of sodium channel blockers which are used asthe active effective ingredients in the ophthalmic compositions of thepresent invention are described as benzothialzole, phenylbenzothialzole, disopyramide, propafenone, flecainide, lorcainide,aprindine, encainide, GEA-968, azure A, pancuronium, N-methylstrychnine,CNS 1237, BW1003C87, BW619C89, U54494A, PD85639, ralitoline, C1953,lifarizine, zonisamide and riluzole.

[0014] The composition may comprise an ophthalmic solution adapted foradministration to the eye of a mammal in the form of intracameralinjection.

[0015] A direct effect on retinal ganglion cells is an importantdiscovery in accordance with the method of the present invention.However, normal electrical excitability of ganglion cells, crucial forvision, will not be compromised.

[0016] Further, a pharmaceutical composition provided in accordance withthe present invention useful for preventing retinal ganglion cell deathassociated with glaucoma with the composition comprising with its activeingredient one or more compounds having non-inactivating sodium channelblocking activity.

[0017] More specifically, the present invention provides a method forpreventing retinal ganglion cell death associated with glaucoma in ananimal of the mammalian species, including humans, which includes thestep of administering to the retinal ganglion cells of the mammal apharmaceutical composition which comprises as its active ingredient oneor more compounds having non-inactivating sodium channel blockingactivity.

BRIEF DESCRIPTION OF THE DRAWINGS

[0018] The advantages and features of the present invention would bebetter understood by the following description when considered inconjunction with the accompanying drawings.

[0019]FIG. 1 is a diagram of the assumed relevant transport mechanismsfor a retinal ganglion cell under normal conditions; and

[0020]FIG. 2 is a diagram of a retinal ganglion cell under ischemicconditions.

DETAILED DESCRIPTION OF THE INVENTION

[0021] While not wishing to be bound by theory, it is believed that thedeath or loss of axons and associated cell bodies comprising the opticnerve is the result of a lethal increase in the intracellularconcentration of calcium ion (Ca⁺²) resulting from an influx of sodiumion (Na⁺) through a non-inactivating sodium ion channel. While studieshave been conducted on rat optic nerve segments (Stys et al, 1995;Waxman, 1995), no application has been made to ganglion cells. There isno expectation of altering a similar sequence of pathological events inretinal cells to prevent death thereof after anoxia based on earlierexperiments on rat optic nerves because it is unclear whether (1) asimilar sequence of events takes place during glaucoma or (2) whethernoninactivating Na channels are present in mammalian retinal ganglioncells, and, if present, the role these channels play in the destructionof retinal ganglion cells that accompanies vision loss associated withglaucoma.

[0022] The procedure in rat retinal ganglion cell is as follows:

[0023] Following depolarization excitable voltage-dependent Na channelsopen for about one millisec and then close. Provided the cell membraneremains depolarized, the channels will not reopen until the membrane isrepolarized towards its resting state. In contrast to normal excitableNa channels, non-inactivating Na channels can be open at normal restingmembrane potentials and can remain open at depolarized potentials. Underpathophysiological conditions such as adenosine triphosphate (ATP)depletion or sustained depolarization Na influx through non-inactivatingNa channels can substantially increase intracellular Na. This increasein intracellular Na causes the electrogenic Na/Ca²⁺ exchanger (Ransom etal, 1993; Stys, 1995, Waxman et al, 1992) which normally operates topromote efflux of Ca²⁺ from the cell to reverse operation with aresulting large increase in intracellular Ca⁺² concentration. The Ca⁺²concentration of the cell may increase from nanomolar to micromolarlevels with the resulting death of said neuronal cell. (Large increasesin intracellular Ca⁺² have been associated with neuronal cell death andprevention of the increase of intracellular Ca⁺² concentration has beenshown to protect neurons of the central nervous system, and rat opticnerve.) In the optic nerve preparation intracellular Ca²⁺ was notmeasured, however, normal cell Ca²⁺ in most cell types including neuronsis approximately 100-200 nanomolar. When Ca²⁺ rises to micromolar levelsit becomes toxic. Exactly what level of Ca²⁺ in optic nerve triggerscell destruction is not known or at least has not been reported.

[0024] Thus, the compounds utilized in accordance with the method of thepresent invention and in the compositions of the present invention aresodium channel blockers which block the non-inactivating sodium ionchannels of the retinal ganglion cells. The sodium channel blockers ofthe present invention prevent the influx of sodium ions into theneuronal cell through the non-activating sodium channel. Preferably thesodium channel blockers of the present invention will selectively blocksaid non-inactivating sodium channels as opposed to voltage-gated sodiumion channels that inactivate rapidly.

[0025] Pharmaceutically acceptable salts of the sodium channel blockerscan also be used in accordance with the present invention. Apharmaceutically acceptable salt may be any salt which retains theactivity of the parent compound and does not impart any deleterious oruntoward effect on the subject to which it is administered and in thecontext in which it is administered.

[0026] Such a salt may be derived from any organic or inorganic acid orbase. The salt may be a mono or polyvalent ion. Of particular interestwhere the acid function is concerned are the inorganic ions, such asalkali ions, e.g. sodium, potassium, etc. Organic amine salts may bemade with amines, particularly ammonium salts such as mono-, di- andtrialkyl amines, e.g. alkyl amines wherein each alkyl group may compriseup to six carbon atoms, or ethanol amines. Salts may also be formed withcaffeine, tromethamine and similar molecules. It is only important thatthe cation of any salt of a sodium channel blocker utilized in thecompositions or methods of this invention be able to block thenon-inactivating sodium channels of the retinal ganglion cell.

[0027] For protecting against retinal ganglion cell damage in amammalian eye, and particularly for prevention of retinal ganglion cellloss in humans exposed to a condition that causes optic neuron loss, theactive compounds (or mixtures or salts thereof) are administered inaccordance with the present invention to the eye admixed with anophthalmically acceptable carrier. Any suitable, e.g., conventional,ophthalmically acceptable carrier may be employed. A carrier isophthalmically acceptable if it has substantially no long term orpermanent detrimental effect on the eye to which it is administered.Examples of ophthalmically acceptable carriers include water (distilledor deionized water), saline and other aqueous media. In accordance withthe invention, the active compounds are preferably soluble in thecarrier which is employed for their administration, so that the activecompounds are administered to the eye in the form of a solution.Alternatively, a suspension of the active compound or compounds (orsalts thereof) in a suitable carrier may also be employed.

[0028] In accordance with the invention the active compounds (ormixtures or salts thereof) are administered in an ophthalmicallyacceptable carrier in sufficient concentration so as to deliver aneffective amount of the active compound or compounds to the optic nervesite of the eye. Preferably, the ophthalmic, therapeutic solutionscontain one or more of the active compounds in a concentration range ofapproximately 0.0001% to approximately 1% (weight by volume) and morepreferably approximately 0.0005% to approximately 0.1% (weight byvolume).

[0029] Any method of administering drugs to the retinal ganglion cellsite of a mammalian eye may be employed to administer, in accordancewith the present invention, the active compound or compounds to the eyeto be treated. By the term “administering” is meant to include thosegeneral systemic drug administration modes, e.g., injection directlyinto the patient's blood vessels, oral administration and the like,which result in the compound or compounds being systemically available.Also, intercameral injection may be utilized to deliver the sodiumchannel blocker to the retinal ganglion cell site. The primary effect onthe mammal resulting from the direct administering of the activecompound or compounds to the mammal's eye is the prevention of opticnerve loss. Preferably, the active useful compound or compounds areapplied topically to the eye or are injected directly into the eye.

[0030] Injection of ophthalmic preparations, for example ocular drops,gels or creams may be used because of ease of application, ease of dosedelivery and fewer systemic side effects, such as cardiovascularhypotension. An exemplary topical ophthalmic formulation is shown belowin Table I. The abbreviation q.s. means a quantity sufficient to effectthe result or to make volume. TABLE I Ingredient Amount (% W/V) ActiveCompound in accordance about 0.0001 to with the invention, about 1Preservative 0-0.10 Vehicle 0-40 Tonicity Adjustor 1-10 Buffer 0.01-10pH Adjustor q.s. pH 4.5-7.5 antioxidant as needed Purified Water asneeded to make 100%

[0031] Various preservatives may be used in the ophthalmic preparationdescribed in Table I above. Preferred preservatives include, but are notlimited to, benzalkonium potassium, chlorobutanol, thimerosal,phenylmercuric acetate, and phenylmercuric nitrate. Likewise, variouspreferred vehicles may be used in such ophthalmic preparation. Thesevehicles include, but are not limited to, polyvinyl alcohol, povidone,hydroxypropyl methyl cellulose, poloxamers, carboxymethyl cellulose andhydroxyethyl cellulose.

[0032] Tonicity adjustors may be added as needed or convenient. Theyinclude, but are not limited to, salts, particularly sodium chloride,potassium chloride, etc., mannitol and glycerin, or any other suitableophthalmically acceptable tonicity adjustor.

[0033] Various buffers and means for adjusting pH may be used so long asthe resulting preparation is ophthalmically acceptable. Accordingly,buffers include but are not limited to, acetate buffers, citratebuffers, phosphate buffers, and borate buffers. Acids or bases may beused to adjust the pH of these formulations as needed.

[0034] In a similar vein, ophthalmically acceptable antioxidantsinclude, but are not limited to, sodium metabisulfite, sodiumthiosulfate, acetylcysteine, butylated hydroxyanisole, and butylatedhydroxytoluene.

[0035] Those skilled in the art will recognize that the frequency ofadministration depends or the precise nature of the active ingredientand its concentration in the ophthalmic formulation.

[0036] Specific examples of sodium channel blockers which are used asthe active effective ingredients in the ophthalmic compositions of thepresent invention are described as benzothialzole, phenylbenzothialzole, disopyramide, propafenone, flecainide, lorcainide,aprindine, encainide, GEA-968, azure A, pancuronium, N-methylstrychnine,CNS 1237, BW1003C87, BW619C89, U54494A, PD85639, ralitoline, C1953,lifarizine, zonisamide and riluzole.

[0037] A sodium channel blocker, in accordance with the presentinvention, may be identified by the methods disclosed in “TheExtracellular Patch Clamp: A method for Resolving Currents ThroughIndividual Open Channels in Biological Membranes”, Neher et al PflugersArchiv V375 pp 219-228 (1978) and “Improved Patch-Clamp Techniques forHigh-Resolution Current Recording from Cells and Cell-Free MembranePatches”, Hamill et al Pflugers Archiv V391 pp 85-100 (1981). Thesereferences are to be incorporated herein in toto for providing a methodfor identifying sodium channel blockers useful in accordance with thepresent invention.

EXAMPLE

[0038]FIG. 1 shows a representation of a retinal ganglion cell 10, undernormal conditions and assumed relevant transport mechanisms 12, 14, 15,18 responsible for maintaining the sodium (Na+) potassium (K⁺) andcalcium (Ca²⁺) gradients and electrical activity of the cell. As shownunder normal conditions ATP levels are adequate and furnish the fuelneeded to drive the Na⁺/K⁺ pump 14 that maintains the K⁺ and Na⁺gradients, keeping intracellular concentrations of K⁺ high and Na⁺ lowrelative to their respective extracellular concentrations. Thevoltage-gated Na⁺ and K⁺ channels 12, 16 provide the currents that makeup the action potential. The electrogenic Na⁺/Ca²⁺ exchanger 18 keepscellular Ca²⁻ levels within the physiological range (nanomolar).

[0039] If, however, ATP levels should drop, due to somepathophysiological insult, the axon will depolarize and the Na⁺/K⁺gradients will collapse over time as a result of Na⁺/K⁺ pump 14inhibition as shown in FIG. 2 for a cell 20 under ischemic conditions.The rise in cellular Na⁺ is mediated by a subset of voltage-gated Na⁺channels that do not inactivate over time. These Na⁺ channels are coined“noninactivating”. The combination of membrane depolarization andintracellular Na⁺ increase is sufficient to drive the Na⁺/Ca²⁺ exchanger18 backwards (see FIG. 2) such that the ganglion cells load with lethallevels of Ca²⁺. It is assumed that this scenario occurs in the retinalganglion cell in glaucoma.

[0040] Accordingly, in accordance with the present invention thefollowing sequence is expected in the presence of a therapeuticconcentration of a Na⁺ channel blocker selective for the noninactivatingtype. First, the Na⁺ channel blocker would have little or no effect onthe normal action potential. This is crucial for normal ganglion cellfunction. Second, it will block the deleterious increase in cell Na⁺ andthe subsequent lethal increase in cell Ca²⁺. Thus, normal ganglion celldysfunction will be minimized and therefore help prevent the loss ofvisual field associated with glaucoma. In addition, blockers ofnoninactivating Na⁺ channels may yield an additional benefit. This isbecause Na⁺ channels are thought to help prevent excitotoxic glutamaterelease which occurs in neuronal tissue during ischemia, hypoxia andother pathological conditions. Excessive extracellular glutamate levelsare neurodestructive and thus may also be involved in glaucomatous opticneuropathy. Thus, Na⁺ overload and excitotoxic increase in extracellularglutamate in accordance with the present invention may be prevented by atherapeutic concentration of one drug, a blocker of noninactivating Na⁺channels.

[0041] In view of the above, it is clear that the scope of the presentinvention should be interpreted solely on the basis of the followingclaims, as such claims are read in light of the disclosure.

What is claimed is:
 1. A method for altering a plausible sequence ofpathological events in retinal ganglion optic cells associated withglaucoma, the sequence including the pathological depolarization ofretinal ganglion cells, an influx of millimolar amounts of sodium vianon-inactivating sodium channels and a subsequent reversal of thesodium/calcium exchanger, mediated by both membrane depolarization andincreased intracellular sodium, causing a toxic buildup of intracellularcalcium, said method comprising blocking of associated non-inactivatingNa⁺ channels in retinal ganglion cells in order to limit sodium/calciumexchange in the retinal ganglion cells and prevent buildup of thecalcium level in the retinal ganglion cells to a lethal level.
 2. Themethod of claim 1 wherein the blocking comprises administering to theretinal ganglion cells a pharmaceutical composition having an activeingredient with non-inactivating sodium channel blocking activity. 3.The method of claim 2 wherein the composition having non-inactivatingsodium channel blocking activity is selected from the group consistingof benzothialzole, riluzole, phenyl benzothialzole and lifarizine. 4.The method of claim 2 wherein the composition comprises an ophthalmicsolution adapted for administration to the eye of a mammal in the formof intracameral injection.
 5. The method of claim 4 wherein aconcentration of the active ingredient in said composition is betweenabout 0.0001 and about 1 percent weight by volume.
 6. A method formaintaining normal intracellular Na⁺ and Ca⁺ in ganglion cells followinga period of anoxia, said method comprising contacting said ganglionneuronal cells with a composition for blocking of non-inactivatingcalcium channel activity in the retinal ganglion neuronal cells.
 7. Themethod of claim 6 wherein the composition for blocking non-inactivatingsodium channels is selected from the group comprising of benzothialzole,riluzole, phenyl benzothiozole and lifarizine.
 8. The method of claim 7wherein the composition comprises an ophthalmic solution adapted foradministration to the eye of a mammal in the form of intracameralinjection.
 9. The method of claim 8 wherein a concentration of theactive ingredient in said composition is between about 0.0001 and about1 percent weight by volume.
 10. A pharmaceutical composition useful forpreventing retinal ganglion cell death, associated with glaucoma, in theeye of a mammal, the composition comprising as its active ingredient oneor more compounds having non-inactivating sodium channel blockingactivity.
 11. The pharmaceutical composition of claim 10 wherein thecompound having non-inactivating sodium channel blocking activity isselected from the group consisting of benzothialzole riluzole,lubelezole, phenyl benzothiozole and lifarizine.
 12. The pharmaceuticalcomposition of claim 10 where the composition is an ophthalmic solution,adapted for administration to the eye of a mammal in the form of anintracameral injection.
 13. The pharmaceutical composition of claim 12wherein the composition contains approximately 0.0001 to 1 percentweight by volume of said compound having non-inactivating sodium channelblocking activity.
 14. A method for preventing retinal ganglion celldeath, associated with glaucoma, in an animal of the mammalian species,including humans, comprising the step of administering to the ganglionoptic nerve of said mammal a pharmaceutical composition which comprisesas its active ingredient one or more compounds having non-inactivatingsodium channel blocking activity.
 15. The method of claim 14 wherein thecompound having non-inactivating sodium channel blocking activity isselected from the group consisting of benzothialzole riluzole, phenylbenzothiozole and lifarizine.
 16. The method of treatment of claim 15wherein the composition is an ophthalmic solution adapted foradministration to the eye of a mammal in the form of intracameralinjection.
 17. The method of treatment of claim 14 wherein in theophthalmic composition the concentration of the compound havingnon-inactivating sodium channel blocking activity is in the range ofapproximately 0.0001 to 1 percent weight by volume.
 18. A method forproviding neuroprotective effect to retinal ganglion cells in the eye ofa human which comprises the step of administering to the human apharmaceutical composition which comprises as its active ingredient oneor more compounds having non-inactivating sodium channel blockingactivity.
 19. The method of claim 18 wherein the compound havingnon-inactivating sodium channel blocking activity is selected from thegroup consisting of benzothialzole riluzole, phenyl benzothiozole andlifarizine.
 20. The method of claim 19 wherein the composition containsapproximately 0.0001 to 1 percent weight by volume of said compoundhaving non-inactivating sodium channel blocking activity.
 21. Apharmaceutical composition useful for preventing retinal ganglion celldeath, associated with glaucoma, in the eye of a mammal, the compositioncomprising as its active ingredient one or more compounds havingnon-inactivating sodium channel blocking activity enabling recovery ofnormal excitability in retinal ganglion cells following a period ofanoxia.
 22. The composition of claim 21 wherein the recovery of normalexcitability following a one hour period of anoxia is at least 30percent.